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Several morphogenetic sequences occur during esophageal development and birth defects occur due to defects in foregut morphogenesis. This work aimed to record the cellular events in the morphogenesis of rabbits' esophageal epithelium. On the 16th day of gestation, the esophageal epithelium varied from stratified ciliated columnar to stratified squamous type. The surface epithelium presented mucous cells with mucigen granules of various sizes occupying their supranuclear cytoplasm. Cytoplasmic vacuolation was evident in all layers of the esophageal epithelium at this age. On the 18th gestational day, some light cells could be detected in the middle portion of the epithelium, while others occupied the whole epithelial length. On the 21st day, mucous cells are more frequently observed at the apical esophageal part as well as at the surface epithelium. Numerous elongated dark cells could be distinguished embedded between the basal cells. On the 24th gestational day the number of the mucous cells reached its peak. Reaching the 30th gestational day, several lamellar bodies, a keratinized layer and mitotic divisions could be demonstrated, and the number of both mucous and dark cells was greatly decreased. Collectively, detection of surface mucous and dark cells together with the non‐cornified surface in some regions of the rabbit esophageal epithelium at the end of gestation ensure a postnatal development to reach the adult epithelium essential to sustain the passage of the harsh raw food. Future immunohistochemical studies are recommended to investigate the components of secretions in mucous cells and functional studies to highlight the dark cells significance. Research Highlights: Esophageal epithelium of fetal rabbit was analyzed by light and transmission microscopy.Surface epithelium presented mucous cells with mucigen granules of various sizes. They reached their maximum number on 24th day then decreased.On the 16th day, cytoplasmic vacuolation was evident in all epithelial layers.On the 21st day, numerous elongated dark cells could be distinguished embedded between the basal cells.Before birth, several lamellar bodies, a keratinized layer and mitotic divisions could be demonstrated, and the number of both mucous and dark cells was greatly decreased. [ABSTRACT FROM AUTHOR]

Lung cancer (LC) represents one of the most prevalent health issues globally and is a leading cause of tumor-related mortality. Despite being one the most attractive compounds of plant origin due to its numerous biological properties, the therapeutic applications of rutin (RUT) are limited by its disadvantageous pharmacokinetics. Thus, the present study aimed to evaluate in vitro the application of two RUT fatty acids bioconjugates, rutin oleate (RUT-O) and rutin linoleate (RUT-L), as potential improved RUT-based chemotherapeutics in non-small cell lung cancer (NSCLC) treatment. The results indicate that both compounds lacked cytotoxic potential in EpiAirway™ tissues at concentrations up to 125 µM. However, only RUT-L exerted anti-tumorigenic activity in NCI-H23 NSCLC cells after 24 h of treatment by reducing cell viability (up to 47%), proliferation, and neutral red uptake, causing cell membrane damage and lactate dehydrogenase (LDH) leakage, affecting cytoskeletal distribution, inducing cytoplasmic vacuolation, and increasing oxidative stress. The cytopathic effects triggered by RUT-L at 100 and 125 µM are indicators of a non-apoptotic cell death pathway that resembles the characteristics of paraptosis. The novel findings of this study stand as a basis for further investigations on the anti-cancer properties of RUT-L and their underlying mechanisms. [ABSTRACT FROM AUTHOR]

Copper alloy sheets have been shown to prevent cryptocaryoniasis. Therefore, we studied the potential efficiency of copper alloy mesh (CAM) in aquaculture tanks to prevent cryptocaryoniasis outbreaks. The effectivenesses of CAM against the tomont stage of Cryptocaryon irritans and in protecting fish from cryptocaryoniasis were tested both in vitro and in vivo. The mortality rate of C. irritans tomonts increased as the contact time with CAM rose and peaked at 70 min (100% of mortality). Morphological changes were observed such as the shrinking of the protoplasm of the treated tomonts, resulting in a larger gap between the cytoplasm and the cyst wall. Mitochondrial dysfunction due to shrinkage in the inner portion, outer and inner mitochondrial membrane damage and cytoplasmic vacuolation was revealed by ultrastructural analysis. The use of CAM effectively preventing reinfection was also provided. In comparison with group B (infected fish without CAM), both groups A (uninfected fish as a control group) and C (infected fish treated with CAM) had a 100% survival rate until the end of the trial. CAM has the same anticryptocaryoniasis effect as copper alloy sheets but is more advantageous due to its lightweight, reduced labor cost and lower purchase cost. It is noticeable that CAM exposure also prevents the excessive accumulation of copper ions in aquaculture sea water. [ABSTRACT FROM AUTHOR]

Glioblastoma multiforme (GBM) is an aggressive brain tumor with high risks of recurrence and mortality. Chemoradiotherapy resistance has been considered a major factor contributing to the extremely poor prognosis of GBM patients. Therefore, there is an urgent need to develop highly effective therapeutic agents. Here, we demonstrate the anti-tumor effect of morusin, a typical prenylated flavonoid, in GBM through in vivo and in vitro models. Morusin showed selective cytotoxicity toward GBM cell lines without harming normal human astrocytes when the concentration was less than 20 µM. Morusin treatment significantly induced apoptosis of GBM cells, accompanied by the activation of endoplasmic reticulum (ER) stress, and the appearance of cytoplasmic vacuolation and autophagosomes in cells. Then, we found the ER stress activation and cytotoxicity of morusin were rescued by ER stress inhibitor 4-PBA. Furthermore, morusin arrested cell cycle at the G1 phase and inhibited cell proliferation of GBM cells through the Akt–mTOR–p70S6K pathway. Dysregulation of ERs and cell cycle in morusin exposed GBM cells were confirmed by RNA-seq analysis. Finally, we demonstrated the combination of morusin and TMZ remarkably enhanced ER stress and displayed a synergistic effect in GBM cells, and suppressed tumor progression in an orthotopic xenograft model. In conclusion, these findings reveal the toxicity of morusin to GBM cells and its ability to enhance drug sensitivity to TMZ, suggesting the potential application value of morusin in the development of therapeutic strategies for human GBM. [ABSTRACT FROM AUTHOR]

Objective: This study aimed to explore the molecular mechanism of cytoplasmic vacuolation caused by BK polyomavirus (BKPyV) and thus search for potential target for drug repurposing. Methods: Morphological features of BK polyomavirus-associated nephropathy (BKPyVAN) were studied under light and electron microscopes. Microarray datasets GSE75693, GSE47199, and GSE72925 were integrated by ComBat, and differentially expressed genes (DEGs) were analyzed using limma. Furthermore, the endoplasmic reticulum (ER)-related genes obtained from GenCLiP 2.0 were intersected with DEGs. GO and KEGG enrichment pathways were performed with intersection genes by R package clusterProfiler. The single-cell RNA sequencing (scRNA-seq) from a BKPyVAN recipient was analyzed with a dataset (GSE140989) downloaded from Gene Expression Omnibus (GEO) as control for gene set variation analysis (GSVA). Immunohistochemistry and electron microscopy of kidney sections from drug-induced ERS mouse models were performed to explore the association of ERS and renal tubular vacuolation. Protein–protein interaction (PPI) network of the intersection genes was constructed to identify hub target. AutoDock was used to screen Food and Drug Administration (FDA)-approved drugs that potentially targeted hub gene. Results: Light and electron microscopes exhibited obvious intranuclear inclusions, vacuoles, and virus particles in BKPyV-infected renal tubular cells. Transcriptome analysis revealed 629 DEGs between samples of BKPyVAN and stable transplanted kidneys, of which 16 were ER-associated genes. GO analysis with the intersection genes illustrated that ERS-related pathways were significantly involved, and KEGG analysis showed a prominent enrichment of MAPK, Toll-like receptor, and chemokine signaling pathways. GSVA analysis of the proximal tubule revealed similar pathways enrichment. An electron microscope image of the kidney from ERS mouse models showed an obvious renal tubular vacuolation with prominent activation of ERS markers verified by immunohistochemistry. Furthermore, DDIT3 was identified as the hub gene based on PPI analysis, and ZINCOOOOO1531009 (Risedronate) was indicated to be a potential drug for DDIT3. Conclusion: ERS was involved in renal tubular cytoplasmic vacuolation in BKPyVAN recipients. Risedronate was screened as a potential drug for BKPyVAN by targeting DDIT3. [ABSTRACT FROM AUTHOR]

From April 2014 to September 2015, 153 piglets from 52 farms in Jeju were diagnosed with porcine epidemic diarrhea (PED). The major PED cases were focused on suckling piglets (144 piglets, 94.1%), particularly in 1-7-day-old piglets. Histopathologically, severe villous atrophy was observed in the small intestine, especially in the jejunum and ileum. The mean villous height to crypt depth ratios of the jejunum and ileum were 1.4:1 and 1.5:1, respectively. The major histopathologic findings of the small intestine were cytoplasmic vacuolation, cuboidalization, squamation, and exfoliation of the mucosal enterocytes in the villi. The cytoplasmic vacuolations in the enterocytes were the most prevalent lesions in the small intestine and were more severe in the ileum than in the jejunum. According to immunohistochemistry methods, the PED virus (PEDV) antigens were presented in the cytoplasms of the enterocytes, and were distributed more prevalently in the ileum than in the jejunum. PEDV antigens were also detected in the colon of 26 piglets (19.5%). Sequence comparison and phylogenetic analysis indicated that 12 PEDV had more than a 98.9% homology with each other. These PEDV strains were highly homologous with the genogroup 2 North American group. [ABSTRACT FROM AUTHOR]

VEXAS (vacuoles, E1 enzyme, X-linked, autoinflammatory, somatic) syndrome is a newly-described adult-onset inflammatory syndrome characterized by vacuoles in myeloid and erythroid precursor cells and somatic mutations affecting methionine-41 (p.Met41) in UBA1. The VEXAS syndrome often overlaps with myelodysplastic syndromes (MDS) with autoimmune disorders (AD). By screening the UBA1 gene sequences derived from MDS patients with AD from our center, we identified one patient with a p.Met41Leu missense mutation in UBA1, who should have been diagnosed as MDS comorbid with VEXAS syndrome. This patient respond poorly to immune suppressive drugs. Patients with MDS and AD who have characteristic vacuoles in myeloid and erythroid precursor cells should be screened for UBA1 mutation, these patients are likely to have VEXAS syndrome and unlikely to improve with immunosuppressive drugs and should be considered for other alternative therapies. [ABSTRACT FROM AUTHOR]

Purpose: Paclitaxel is an anticancer drug for the treatment of non-small cell lung cancer (NSCLC). However, drug-resistance remains a major problem. Honokiol is a natural component which has been found to exhibit anti-tumor activity. Paclitaxel and honokiol have been reported to be able to induce paraptosis. The aim of this study was to investigate whether honokiol can reverse paclitaxel resistance by inducing paraptosis in NSCLC cells. Methods: NSCLC cell lines H1650 (paclitaxel-sensitive), H1299 and H1650/PTX (intrinsic and acquired paclitaxel-resistant, respectively) were used to assess the cytotoxic effects of paclitaxel and honokiol. Light and transmission electron microscopy were performed to detect cytoplasmic vacuolation. In vitro cell viability and clonogenic survival assays, as well as in vivo xenograft assays were conducted to test synergistic killing effects of paclitaxel and honokiol on NSCLC cells. Western blotting, flow cytometry and immunofluorescence were performed to evaluate paraptosis-regulating mechanisms. Results: We found that combination treatment with paclitaxel and honokiol synergistically killed H1650, H1299 and H1650/PTX cells by inducing paraptosis, which is characterized by cytoplasmic vacuolation. Moreover, paclitaxel/honokiol treatment resulted in a significant growth delay in H1299 xenograft tumors that showed extensive cytoplasmic vacuolation. Mechanistically, proteasomal inhibition-mediated endoplasmic reticulum (ER) stress and unfolded protein responses leading to ER dilation, and the disruption of intracellular Ca2+ homeostasis and mitochondrial Ca2+ overload resulting in mitochondrial disfunction, were found to be involved in paclitaxel/honokiol-induced paraptosis. Cellular protein light chain 3 (LC3) may play an important role in paclitaxel/honokiol induced cytoplasmic vacuolation and NSCLC cell death. Conclusions: Combination of honokiol and paclitaxel may represent a novel strategy for the treatment of paclitaxel-resistant NSCLC. [ABSTRACT FROM AUTHOR]

Chalcones are biologically active class of compounds, known for their anticancer activities. Here we show for the first time that out of the six synthetic derivatives of chalcone tested, 2′‐hydroxy‐retrochalcone (HRC) was the most effective in inducing extensive cytoplasmic vacuolation mediated death called paraptosis in malignant breast and cervical cancer cells. The cell death by HRC is found to be nonapoptotic in nature due to the absence of DNA fragmentation, PARP cleavage, and phosphatidylserine externalization. It was also found to be nonautophagic as there was an increase in the levels of autophagic markers LC3I, LC3II and p62. Immunofluorescence with the endoplasmic reticulum (ER) marker protein calreticulin showed that the cytoplasmic vacuoles formed were derived from the ER. This ER dilation was due to ER stress as evidenced from the increase in polyubiquitinated proteins, Bip and CHOP. Docking studies revealed that HRC could bind to the Thr1 residue on the active site of the chymotrypsin‐like subunit of the proteasome. The inhibition of proteasomal activity was further confirmed by the fluorescence based assay of the chymotrypsin‐like subunit of the 26S proteasome. The cell death by HRC was also triggered by the collapse of mitochondrial membrane potential and depletion of ATP. Pretreatment with thiol antioxidants and cycloheximide were able to inhibit this programmed cell death. Thus our data suggest that HRC can effectively kill cancer cells via paraptosis, an alternative death pathway and can be a potential lead molecule for anticancer therapy. [ABSTRACT FROM AUTHOR]

Silver nanoparticles (Ag NPs) are invested in various sectors and are becoming more persistent in our ambient environment with potential risk on our health and the ecosystems. The current study aims to investigate the histological, histochemical and ultrastructural hepatic changes that might be induced by 10 nm silver nanomaterials. Male mice (BALB/C) were exposed for 35 injections of daily dose of 10 nm Ag NPs (2 mg/kg). Liver tissues were subjected to examination by light and electron microscopy for histological, histochemical and ultrastructural alterations. Exposure to Ag NPs induced Kupffer cells hyperplasia, sinusoidal dilatation, apoptosis, ground glass hepatocytes appearance, nuclear changes, inflammatory cells infiltration, hepatocytes degeneration and necrosis. In addition, 10 nm Ag NPs induced histochemical alterations mainly glycogen depletion with no hemosiderin precipitation. Moreover, these nanomaterials exhibited ultrastructure alterations including mitochondrial swelling and cristolysis, cytoplasmic vacuolation, apoptosis, multilammellar myelin figures formation and endoplasmic destruction and reduction. The findings revealed that Ag NPs can induce alterations in the hepatic tissues, the chemical components of the hepatocytes and in the ultrastructure of the liver. One may also conclude that small size Ag NPs, which are increasingly used in human products could cause various toxigenic responses to all hepatic tissue components. [ABSTRACT FROM AUTHOR]

We investigated the antitumor activity of daucusol (DS) derived from Daucus carota L. in PC-3, A549 and HeLa cell lines by the MTT assay. Optical microscopy revealed that exposure of PC-3 cells to DS resulted in cytoplasmic vacuolation. Flow cytometry analysis of the phase of the cell cycle did not reveal a sub-G1 peak and no caspase-dependent activation was observed after DS treatment. The levels of endoplasmic reticulum (ER) stress biomarkers, LC3B-II and ubiquitinated proteins were increased. It was also observed that oxidative stress played an important role in the activation of the cytoplasmic vacuolation-mediated cell-death pathway. In vivo, DS inhibited tumor growth in nude mice by 39.13% compared to the vehicle. Protein expression in the tumor tissue was consistent with their expression in vitro. Our findings indicate that DS induced cytoplasmic vacuolation-mediated death in PC-3 cells by triggering oxidative stress and suggest that targeting this pathway could serve as a novel therapeutic approach for prostate cancer. [ABSTRACT FROM AUTHOR]

The two evolutionarily conserved mammalian lipid kinases Vps34 and PIKfyve are involved in an important physiological relationship, whereby the former produces phosphatidylinositol (PtdIns) 3P that is used as a substrate for PtdIns(3,5)P2 synthesis by the latter. Reduced production of PtdIns(3,5)P2 in proliferating mammalian cells is phenotypically manifested by the formation of multiple translucent cytoplasmic vacuoles, readily rescued upon exogenous delivery of PtdIns(3,5)P2 or overproduction of PIKfyve. Although the aberrant vacuolation phenomenon has been frequently used as a sensitive functional measure of localized PtdIns(3,5)P2 reduction, cellular factors governing the appearance of cytoplasmic vacuoles under PtdIns3P-PtdIns(3,5)P2 loss remain elusive. To gain further mechanistic insight about the vacuolation process following PtdIns(3,5)P2 reduction, in this study we sought for cellular mechanisms required for manifestation of the aberrant endomembrane vacuoles triggered by PIKfyve or Vps34 dysfunction. The latter was achieved by various means such as pharmacological inhibition, gene disruption, or dominant-interference in several proliferating mammalian cell types. We report here that inhibition of V-ATPase with bafilomycin A1 as well as inactivation of the GTP-GDP cycle of Rab5a GTPase phenotypically rescued or completely precluded the cytoplasmic vacuolization despite the continued presence of inactivated PIKfyve or Vps34. Bafilomycin A1 also restored the aberrant EEA1-positive endosomes, enlarged upon short PIKfyve inhibition with YM201636. Together, our work identifies for the first time that factors such as active V-ATPase or functional Rab5a cycle are acting coincidentally with the PtdIns(3,5)P2 reduction in triggering formation of aberrant cytoplasmic vacuoles under PIKfyve or Vps34 dysfunction. [ABSTRACT FROM AUTHOR]

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Thiol reactive cyclopentenone prostaglandin, 15-deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2), induced a novel, nonapoptotic and microtubule-associated protein 1 light chain 3 (MAP1 LC3) dependent but nonautophagic form of cell death in colon, breast and prostate cancer cell lines, characterized by extensive cytoplasmic vacuolation with dilatation of endoplasmic reticulum (ER). Disruption of sulfhydryl homeostasis, which resulted in ER stress, accumulation of ubiquitinated proteins and subsequent ER dilation, contributed to peroxisome proliferator-activated receptor γ (PPARγ)-independent cell death by 15d-PGJ2. Absence of intracellular organelles in these vacuoles, shown by electron microscopy and unique fragmentation of lamin B, suggested this form of cell death to be different from autophagy and apoptosis. Cell death induced by 15d-PGJ2 is prevented by cycloheximide and actinomycin D, suggesting a requirement of new protein synthesis for death with cytoplasmic vacuolation. Here, we report for the first time that upregulation and processing of autophagy marker LC3 is an important event in nonautophagic cytoplasmic vacuolation and cell death. Notably, knockdown of LC3 conferred significant protection against 15d-PGJ2-induced cytoplasmic vacuolation and cell death, suggesting a novel role of LC3 in a death process other than autophagy.Oncogene (2009) 28, 2556–2568; doi:10.1038/onc.2009.118; published online 18 May 2009 [ABSTRACT FROM AUTHOR]

This study evaluated the synergistic anti-cancer potential of cannabinoid combinations across the MDA-MB-231 and MCF-7 human breast cancer cell lines. Cannabinoids were combined and their synergistic interactions were evaluated using median effect analysis. The most promising cannabinoid combination (C6) consisted of tetrahydrocannabinol, cannabigerol (CBG), cannabinol (CBN), and cannabidiol (CBD), and displayed favorable dose reduction indices and limited cytotoxicity against the non-cancerous breast cell line, MCF-10A. C6 exerted its effects in the MCF-7 cell line by inducing cell cycle arrest in the G2 phase, followed by the induction of apoptosis. Morphological observations indicated the induction of cytoplasmic vacuolation, with further investigation suggesting that the vacuole membrane was derived from the endoplasmic reticulum. In addition, lipid accumulation, increased lysosome size, and significant increases in the endoplasmic reticulum chaperone protein glucose-regulated protein 78 (GRP78) expression were also observed. The selectivity and ability of cannabinoids to halt cancer cell proliferation via pathways resembling apoptosis, autophagy, and paraptosis shows promise for cannabinoid use in standardized breast cancer treatment. [ABSTRACT FROM AUTHOR]